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Title:
METHOD FOR PRODUCING AAV GENE DRUGS ON BASIS OF INDUCIBLE INSECT CELLS
Document Type and Number:
WIPO Patent Application WO/2022/160512
Kind Code:
A1
Abstract:
A method for producing AAV gene drugs on the basis of inducible insect cells. The method comprises: (1) respectively constructing a Rep gene plasmid, a Cap gene plasmid and a target gene plasmid for packaging an adeno-associated virus, wherein the Rep gene plasmid and the Cap gene plasmid contain a late promoter; (2) integrating the three constructed plasmids into an Sf9 insect cell genome to obtain a stable transfected cell strain capable of producing AAV; and (3) performing mass amplification, and culturing the stable transfected cell strain. When the AAV needs to be produced, a baculovirus carrying no exogenous gene is used to infect the stable transfected cell strain, so as to induce the late promoter in the stable transfected cell strain to activate and drive the expression of the Rep gene and Cap gene, and package and produce AAV. The method does not need to use a baculovirus carrying an exogenous gene, so that the problem of a loss of exogenous genes caused by mass replication and amplification of the baculovirus is avoided, which is conducive to obtain a stable AAV product, the steps for producing AAV are further simplified, the production efficiency is improved, the large-scale efficient production of the AAV is achieved, and the cost is reduced.

Inventors:
DONG WENJI (CN)
LIU ZIJIN (CN)
ZHAO ZHONGLIANG (CN)
CAO FAN (CN)
CHENG MOBIN (CN)
ZHANG YANJUN (CN)
Application Number:
PCT/CN2021/093906
Publication Date:
August 04, 2022
Filing Date:
May 14, 2021
Export Citation:
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Assignee:
GENMEDICN BIOPHARMA INC (CN)
International Classes:
C12N15/864; C07K14/47; C12N5/10; C12N7/01; C12N9/48; C12N15/12; C12N15/35; C12N15/55; C12N15/866
Domestic Patent References:
WO2017184879A12017-10-26
Foreign References:
CN112852880A2021-05-28
CN106916793A2017-07-04
CN106987603A2017-07-28
CN106085972A2016-11-09
Other References:
YANG WU, LIANGYU JIANG, HAO GENG, TIAN YANG, ZENGPENG HAN, XIAOBING HE, KUNZHANG LIN, FUQIANG XU: "A Recombinant Baculovirus Efficiently Generates Recombinant Adeno-Associated Virus Vectors in Cultured Insect Cells and Larvae", MOLECULAR THERAPY- METHODS & CLINICAL DEVELOPMENT, NATURE PUBLISHING GROUP, GB, vol. 10, 1 September 2018 (2018-09-01), GB , pages 38 - 47, XP055681042, ISSN: 2329-0501, DOI: 10.1016/j.omtm.2018.05.005
LEI, QIUGANG: "Establishment of Production System of Recombinant Adeno-associated Viruses Based on Baculoviruses", JOURNAL OF ZHEJIANG SCI-TECH UNIVERSITY, vol. 27, no. 5, 1 September 2010 (2010-09-01), pages 1ยง - 6, XP055953564, ISSN: 1673-3851
WU YANG, MEI TING, JIANG LIANGYU, HAN ZENGPENG, DONG RUPING, YANG TIAN, XU FUQIANG: "Development of Versatile and Flexible Sf9 Packaging Cell Line-Dependent OneBac System for Large-Scale Recombinant Adeno-Associated Virus Production", HUMAN GENE THERAPY METHODS, vol. 30, no. 5, 1 October 2019 (2019-10-01), pages 172 - 183, XP055845975, ISSN: 1946-6536, DOI: 10.1089/hgtb.2019.123
MARIO MIETZSCH, HENRIK HERING, EVA-MARIA HAMMER, MAVIS AGBANDJE-MCKENNA, SERGEI ZOLOTUKHIN, REGINE HEILBRONN: "OneBac 2.0: Sf 9 Cell Lines for Production of AAV1, AAV2, and AAV8 Vectors with Minimal Encapsidation of Foreign DNA", HUMAN GENE THERAPY METHODS, vol. 28, no. 1, 1 February 2017 (2017-02-01), pages 15 - 22, XP055417034, ISSN: 1946-6536, DOI: 10.1089/hgtb.2016.164
MICHAEL F. NASO, BRIAN TOMKOWICZ, WILLIAM L. PERRY, WILLIAM R. STROHL: "Adeno-Associated Virus (AAV) as a Vector for Gene Therapy", BIODRUGS, ADIS INTERNATIONAL LTD., NZ, vol. 31, no. 4, 1 August 2017 (2017-08-01), NZ , pages 317 - 334, XP055547503, ISSN: 1173-8804, DOI: 10.1007/s40259-017-0234-5
Attorney, Agent or Firm:
BEIJING EASY JOY SOON IP AGENT LTD (CN)
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